Project Abstract Pancreatic Ductal Adenocarcinoma (PDA) is the most commonly diagnosed form (~90%) of pancreatic cancer with a dismal 5-year survival rate of ~8%. PDA features a prominent pro-tumorigenic microenvironment rich in immunosuppressive cells that inhibit functions of anti-tumor immunity. Current immunotherapeutic approaches have been unsuccessful in improving PDA patient outcomes, leaving a need for a better understanding of the immunomodulatory signaling mechanisms within pancreatic tumor microenvironment. We have identified infiltrating regulatory B cells as being key contributors to pancreatic tumorigenesis through their expression of the regulatory cytokine IL-35, which directly inhibits anti-tumor immune responses. The mechanisms that cause B cells to produce IL-35 in cancer and promote pancreatic tumor growth are not known. Using a mouse model harboring a monoclonal fixed B cell receptor (BCR), or mice with a B cell-specific deletion of the Toll-Like Receptor (TLR) adaptor protein MyD88, I found that orthotopically implanted pancreatic tumors were drastically reduced in size as compared to WT mice. I have also conducted in vitro analysis of TLR and BCR activation and found that co-stimulation of endosomal TLRs and the BCR leads to a robust increase of IL-35 expression in B cells. Based on my preliminary data, I hypothesize that cancer-driven upregulation of IL-35 in B cells is dependent on crosstalk between BCR and endosomal TLRs, which promotes pancreatic tumor growth through immunosuppression. I propose two specific aims to test my hypothesis. In Aim 1, I will determine how BCR and endosomal TLR signaling via Bruton's tyrosine kinase (BTK) contributes to expression of IL-35 expression in B cells, using analysis of signaling pathways in primary regulatory B cells and established B cell lines. In Aim 2, I will investigate how activation of both BCR and TLR signaling in B cells promotes PDA tumor growth in vivo. To accomplish this task, I will analyze mouse models expressing a fixed BCR specificity with or without specific antigen exposure, mouse models lacking MyD88 signaling in B cells, as well as a cross of the two models. Our proposed research will provide an understanding of a previously uncharacterized facet of B cell-mediated function in PDA and use state-of-the-art PDA murine models to test strategies that block immune suppressive pathways. Ultimately, I anticipate my findings will reveal a targetable mechanism to inhibit B cell-mediated immunosuppression in pancreatic tumors and provide a multi-faceted training experience to help advance my scientific career.